Cell transfection apparatus and method

ABSTRACT

An apparatus and associated method provides for the application of a cell treatment agent, such as genetic material or drugs to be inserted within the cells of a patient in vivo. The apparatus may be a catheter arrangement with various embodiments for applying heat to a patient&#39;s cells in vivo in order to improve transfection efficiency or application efficiency. Laser beams may be applied directly to the cells. Alternately, the cells may be heated by electrical heating, chemical heating, radio frequency heating, microwave heating, infrared heating, ultrasound heating, or indirect laser heating. Further, the treatment agent may be heated prior to its application to the patient such that the treatment agent heats the cells of the patient.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional application of co-pending U.S.application Ser. No. 08/773,509, filed Dec. 23, 1996, now U.S. Pat. No.6,071,276 issued Jun. 6, 2000. That latter application was in turn adivisional application of co-pending U.S. application Ser. No.08/276,324, filed Jul. 18, 1994, now U.S. Pat. No. 5,586,982 issued Dec.24, 1996. That 08/276,324 application in turn was a continuation-in-partof co-pending U.S. Ser. No. 08/053,206 filed Apr. 28, 1993, now U.S.Pat. No. 5,330,467 issued Jul. 19, 1994, which in turn was acontinuation of Ser. No. 07/866,473 filed Apr. 10, 1992, now U.S. Pat.No. 5,246,437 issued Sep. 11, 1993. All of those prior applications werefiled in the name of the present inventor. Those last two mentionedapplications are both entitled “CELL TREATMENT METHOD AND APPARATUS” andboth are hereby incorporated by reference.

BACKGROUND OF THE INVENTION

This invention relates to a cell treatment apparatus and method. Morespecifically, this invention relates to a treatment apparatus and methodfor transfecting a patient's cells in vivo.

Various techniques have been used, at least experimentally, fortransfection of cells (i.e., insertion of new genetic material into theDNA structure of cells).

One technique for transfection of cells has used laser poration. Thisapproach has been performed in vitro using a laser beam to porate asingle cell at a time under a specially adapted microscope. Themicroscope allows the direct puncture of the cell membrane in thepresence of the gene. Specifically, an operator directs the laser beamtowards an individual cell and the puncture of the cell membrane allowsgenetic material on the same side as the cell to enter into the cell.This approach is labor intensive and not practical to use in vivo. Thislaser poration technique is described in the article by Tao et al.entitled “Direct Gene Transfer Into Human Cultured Cells Facilitated ByLaser Micropuncture of Cell Membrane” in the Proceedings of the NationalAcademy of Science in 1987; 84:4180-4184.

Other approaches to transfection of cells have included chemical methodsor electrical poration used in a cell culture, but such methods have notbeen readily applicable in vivo. In other words, such methods may allowtreatment of cells which have been removed from the patient, but do notallow treatment of cells remaining with the patient (human or animal).

The Nabel et al. article entitled “Site-Specific Gene Expression in Vivoby Direct Gene Transfer Into the Arterial Wall” in Science in 1990;249(4974):1285-1288 discloses a technique for transfecting genes in vivowhich has been used in the arteries of pigs. This technique uses acatheter with a dual balloon system at the tip of the catheter. The twoballoons are inflated to create a temporary chamber which allows theexposure of the arterial wall to a viral transporting agent in solution.This has been used successfully to transfect the arterial wall with aDNA-plasmid having a viral carrier. However, this double balloon methodrequires 30 minutes to bathe the arterial wall with the DNA-plasmid tobe effective. This is not feasible in certain applications such as inthe coronary circulation. Moreover, the time required for such atechnique to work may pose severe problems even at other locationswithin the arteries of an animal or human.

The Lim et al. article entitled “Direct In Vivo Gene Transfer Into theCoronary and Peripheral Vasculatures of the Intact Dog” appearing inCirculation, volume 83, no. Jun. 6, 1991, pages 2007-2011, discloses atechnique where endothelial cells are removed from the test animal andthen transfected prior to reintroduction into the animal. In addition tothat in vitro technique, the article describes in vivo transfection ofarteries of dogs using catheters placed in peripheral vessels of thedogs. Proximal and distal lumens of the vessels were occluded withremovable ligatures. In somewhat similar fashion to the dual balloonsystem, a temporary chamber is established and a transfection solutionis supplied into that temporary chamber within the vessels of theanimal. The article describes allowing the transfection solution toremain in the vessel for one hour.

In the two above incorporated by reference applications, the presentinventor has disclosed cell treatment (more specifically celltransfection) of a patient's cells in vivo by use of a laser catheter.No admission is made or intended that these prior applications of thepresent inventor are necessarily prior art to the present application.However, it is noted that the present inventor has discovered the use ofvarious additional techniques for in vivo transfection of a patient'scells. As used herein, in vivo shall refer to treatment of a patient'scells without removing the cells from the patient. Thus, in vivotreatment involves treatment of cells within or on the patient.

OBJECTS AND SUMMARY OF THE INVENTION

Accordingly, it is a primary object of the present invention to providea new and improved cell transfection apparatus and method.

A more specific object of the present invention is to provide for highlyefficient cell transfection in vivo.

A further object of the present invention is to provide for transfectionof cells in vivo relatively quickly (i.e., the cells in vivo need not beexposed for such long periods that lengthy disruptions, such as blockingof artery flow, are required).

The above and other objects which will become more apparent as thedescription proceeds are realized by an apparatus for cell transfectionincluding an instrument with a housing having a wall with at least onehole therein. A heater is operatively connected to the instrument so asto apply heat to cells of a patient in vivo. A source of treatment agentincluding a DNA plasmid is operatively connected to the instrument so asto apply treatment agent to the heated cells by way of a treatmentchannel inside the instrument and extending to the hole. The treatmentagent is operable to transfect the heated cells.

The heater, which may also be called a heating means, may be realized byvarious alternate constructions. In a first embodiment, the heaterincludes an optical fiber within the instrument for applying laserenergy by way of the hole to heat the cells. In a second embodiment, theheater includes an optical fiber within the instrument for heating anopaque portion of the instrument such that the opaque portion in turnheats the cells. In a third embodiment, the heater is an RF source whichheats the cells by application of radio frequency energy. In a fourthembodiment, the heater is a microwave source which heats the cells byapplication of microwave energy. In a fifth embodiment, the heater is anelectrical heater mounted to the housing. In a sixth embodiment, theheater is chemical heating material adjacent the hole. In a seventhembodiment, the instrument has a distal portion adjacent the hole and aproximal portion remote from the hole and the heater is adjacent theproximal portion for heating the cells by heating treatment fluid remotefrom the distal portion. Other embodiments use infrared or ultrasound inorder to heat patient cells.

Regardless of the heater arrangement, the instrument is preferably acatheter for insertion in a patient. The catheter further includes aflushing solution channel terminating in a flushing solution exit forapplying flushing solution to a treatment site in a patient. Thecatheter includes a balloon mounted thereon and a balloon channelconnected to the balloon for inflating the balloon, the balloon servingto occlude a body passage when the catheter is used for celltransfection.

The method of transfecting cells of a patient in vivo includes applyingheat to the cells of the patient in vivo and providing a treatment agentincluding a DNA plasmid to the heated cells such that the heated cellsare transfected. The application of heat is accomplished from one ormore of the steps selected from the group including: application oflaser energy to the cells, application of laser energy to heat an opaqueportion of a part of an apparatus in thermal transfer position relativeto the patient such that the apparatus in turn heats the cells, theapplication of radio frequency energy to the cells, application ofmicrowave energy to the cells, electrically heating the cells,chemically heating the cells, heating-the cells by infrared energy,heating the cells by ultrasound energy, and heating the treatment agentprior to its insertion in the patient such that the treatment agentheats the cells.

Preferably, the method further includes inserting a catheter into thepatient, the catheter having a treatment agent channel, and wherein theheat is applied by way of the catheter and the treatment agent isprovided by way of the treatment agent channel.

In one technique according to the present invention, heat is appliedusing application of laser energy exiting from the catheter in the formof a plurality of distinct beams including at least first and secondbeams which cause porations in the cells, the first beam applied to afirst one of the cells when the second beam is applied to a second oneof the cells.

Heat may be applied by use of laser energy exiting from the catheter inthe form of a beam leaving the catheter with a width of less than 100microns.

Heat may be applied by use of application of laser energy exiting fromopenings on a side of the catheter in the form of a plurality of beams,each leaving the catheter with a width of less than 100 microns, theplurality of beams causing porations in the cells, and the treatmentagent is forced out the openings to follow multiple paths correspondingto the beams.

Heat may be applied by application of laser energy to heat an opaqueportion of a part of the catheter in thermal transfer position relativeto the patient such that the catheter in turn heats the cells.

BRIEF DESCRIPTION OF THE DRAWINGS

The above and other features of the present invention will be morereadily understood when the following detailed description is consideredin conjunction with the accompanying drawings wherein like charactersrepresent like parts throughout the several views and in which:

FIG. 1 shows a side view of a treatment catheter arrangement accordingto the present invention;

FIG. 2 shows a side view, with parts in cross section, of the treatmentcatheter or catheter assembly of FIG. 1;

FIG. 2A shows a simplified end view of the laser catheter;

FIG. 3 shows an end view of a tip of the arrangement of FIG. 2;

FIG. 4 shows a screen or window portion used to provide a plurality ofvery small beams of laser energy from a single larger beam of laserenergy;

FIG. 5 shows a schematic view of a laser beam genetic code materialbeing applied to different cells;

FIG. 6 shows a side view of a second embodiment catheter according tothe present invention;

FIG. 7 shows a simplified side view with parts in cross-section of adistal end of a third embodiment catheter;

FIG. 8 shows a simplified side view with parts in cross-section of adistal end of a fourth embodiment catheter;

FIG. 9 shows a simplified side view with parts in cross-section of adistal end of a fifth embodiment catheter;

FIG. 10 shows a simplified side view with parts in cross-section of adistal end of a sixth embodiment catheter;

FIG. 11 shows a simplified side view with parts in cross-section of adistal end of a seventh embodiment catheter;

FIG. 12 shows a simplified side view with parts in cross-section of adistal end of a eighth embodiment catheter;

FIG. 13 shows a simplified side view with parts in cross-section of adistal end of a ninth embodiment catheter with parts of a patient;

FIG. 14 shows a simplified front view schematic to illustrate staggeringof various fibers in yet another embodiment;

FIG. 15 is a side view with parts in cross section of a catheter usingthe staggering feature of FIG. 14;

FIG. 16 is a cross section with parts removed for simplicity taken alongline 16—16 of FIG. 15;

FIG. 17 shows a perspective of an external end of a fiber optic bundlecorresponding to the FIG. 15 combined with a schematic illustrating howlaser energy is applied to the bundle;

FIG. 18 is a side view with parts in cross section of a catheter using areflector;

FIG. 19 is a detailed side view of the reflector and optical fibers ofFIG. 18; and

FIG. 20 is a view of the face of an alternate reflector used to split arelatively broad beam into a plurality of small width beams.

DETAILED DESCRIPTION

Turning initially to FIG. 1, a brief overview of the preferredembodiment of the present invention will be given. A treatment catheter10, which might also be called a catheter assembly, includes aninsertion catheter 12 and a laser catheter 14. The insertion catheter 12has a balloon 16 mounted thereon for blocking blood flow in an artery,while the laser catheter 14 is applying laser energy 18 to porate cells(not shown) at the same time as genetic code material (not separatelyshown) is proceeding along the same paths as the laser energy 18. Aguide wire 20 is used to guide the catheters to their intended locationsuch that the laser energy 18 and associated genetic material may beapplied to the proper site within the patient.

A balloon syringe 22 controls the balloon 16 in known fashion by way ofa Y connection 24, which may be constructed in known fashion. Saline orother flush may be provided to the insertion catheter 12 by way of Yconnector 26 having entry tube 28 for those purposes. The laser or innercatheter 14 is visible as it extends out the back of connector 26towards the Y connector 30 having tube 32 for entry of genetic materialor a drug to be inserted in the patient. A pump (not shown) or otherarrangement may be connected to tube 32 to supply genetic material or adrug under pressure to inner catheter 14. The Y connector 30 isconstructed in known fashion to merge the guide wire 20 (which proceedsfrom the luer lock or hemostatic Y-connector 34 and tube 36), opticalfiber 38 (operably connected to laser 40 and proceeding through fiberconnector 42 and tube 44), and the material inserted into the entry tube32. Accordingly, the inner or laser catheter 14 proceeding out the leftside of connector 30 includes the guide wire 20, the optical fiber(fiber optic element) 38, and any material supplied to the tube 32.

A feedback loop uses a temperature sensor 81, comparator 81C, and powersupply 82P. The operation of these components control the temperature offluid in a manner as discussed in more detail relative to the FIG. 8embodiment.

Turning now to FIG. 2, the details of the tip of the catheterarrangement of FIG. 1 will be explained. The insertion of flush catheter12, which may be made of a common catheter material such as Teflon typematerial, is generally a hollow cylinder having a balloon controlchannel 46 separated from the main hollow part of the tubular catheter12 and used to control the balloon 16 in known fashion by way of an exit46E for the channel 46. A flushing solution channel 48, which isseparate and distinct from the balloon channel 46, is disposed withinthe main hollow part of catheter 12. More specifically, the channel 48may extend circumferentially in a ring just inside the wall of catheter12 and outside of the inner or laser catheter 14. The channel 48proceeds to a circumferential flush exit 48E.

The inner or laser catheter 14 has a generally cylindrical tube 50 ofcommon flexible material used for catheters. Inside of the tube 50 is aguide wire tube 52 for slidably receiving the guide wire 20 therein. Thetube 52, would be secured to one side of the tube 50 as best understoodby momentarily turning to the end view of laser catheter 14 in FIG. 2A.Instead of having the tube 52 be distinct from tube 50, tube 52 mightsimply be a lengthwise extending pocket in tube 50 extruded at the sametime as tube 50.

Turning back to FIG. 2, the optical fiber 38 extends inside of tube 50and outside of tube 52 and extends into a connecting tube 54, which ispreferably about 4 millimeters long and made of metal. The guide wiretube 52 and the connecting tube 54 would be glued, have barbs (notshown) to grip tube 50, or otherwise fixed in position (possibly simplyby friction) within the tube 50. The connecting tube 54 secures a tip 56to the tube 50. Specifically, the connecting tube 54 may be glued,friction-fit, snap fit using a ledge ring (not shown), or otherwisefixed to a cylindrical bore 56B within the tip 56. A second bore orcylindrical hole 56R (turn momentarily to FIG. 3) extends lengthwise inthe tip 56. The tip 56, which is preferably made of surgical steel andis 8.7 millimeters long in the preferred embodiment, has a tapered end58 which may be made of Teflon or other surgical materials commonly usedin catheters. The tapered portion 58 would be hollow or otherwise allowfor passage of guide wire 20 therethrough.

The optical fiber 38 proceeds through connecting tube 54 to a glass hood60. Since the optical fiber 38 has a smaller outside diameter than theinner diameter of the hollow cylindrical connecting tube 54, an annularpassage 54P is provided therebetween to allow fluid flow of treatmentagent from an annular channel 53 into bore 56B. The optical fiber 38 issecured to the glass hood 60 by way of epoxy 62 applied after heatingand creating a vacuum within the hood 60. The hood 60 preferably has asquare cross section to fit within a square cross section cavity 60C(also refer momentarily to FIG. 3) which extends out from the circularbore or cavity 56B. Accordingly, the glass hood 60 may be adhered,friction-fit, or otherwise fixed within cavity 60C to prevent relativeangular movement between the optical fiber 38 and the tip 56. Theoptical fiber 38 may have a portion of its cladding removed at itsnarrow portion 38N adjacent its end. The optical fiber 38 has a tip 38Tto cause any laser beam to be directed sideways out a window portion 56Win the side of the wall of tip 56. More details of the construction oftip 38T and glass hood 60 may be obtained from U.S. Pat. No. 5,061,265,invented by the present inventor together with Stephan E. Friedl, issuedon Oct. 29, 1991, and hereby incorporated by reference. Generally, thetip 38T is made into a prism using techniques described in that priorpatent so as to deflect all, or substantially all, of the laser energyout the window portion 56W.

Continuing to view FIG. 2, but also referring to the enlarged view ofthe window portion 56W of tip 56 appearing in FIG. 4, the window portion56W is essentially a laser screen having a plurality of very small holes56H through which laser micro-beams 64 may pass. Thus, the windowportion 56W may be considered as a grating portion having grating means,the grating means constituted by the holes or openings 56H and thematerial in between the holes 56H. The holes 56H would be distributedthroughout an area of between one square millimeter and three squaremillimeters. Preferably, the holes are distributed evenly over acircular area of two square millimeters which would correspond to thewidth of the beam exiting from the tip 38T. Each of the holes 56H wouldbe less than 200 microns. More specifically, the holes would be below100 microns in diameter such that the micro-beams 64 would have acorresponding width as they leave the tip 56 of catheter 14. Mostspecifically, the holes would be 50 to 100 microns in diameter toprovide beams of the same size. The holes 56H may be made by using anexcimer laser or electrodischarge machine. The microbeams 64 wouldpreferably have a diameter of 50 to 75 microns. As apparent from FIGS. 2and 4, there are at least three beams which go in the same generaldirection. More specifically, at least 12 beams; corresponding to atleast 12 holes, proceed out one side of the tip 56. As will beappreciated, there is a one-to-one correspondence between the beams 64and the holes 56H. The beams 64 are distinct (i.e., meaning distinctwhere they leave the tip 56), but each of the beams 64 diverges somewhatbecause of the properties of the prism tip 38T of the optical fiber 38.

The openings 56H also allow passage of treatment agent which is suppliedvia treatment agent channel 53 (FIG. 2) in between fiber 38 and tube 50,connecting tube 54 (i.e., between tube 54 and fiber 38) and bore 56B.

Operation

Having described the structural features of the present invention, themethod according to the present invention will now be described.

Although the present invention has applicability to providing treatmentvery efficiently on a cellular level at various sites on a patient'sbody or in a patient's body, the specifics of the structure which hasbeen described is best suited for applying treatment to the walls of anartery and the explanation which follows will emphasize such anapplication of the invention.

A patient having an artery, such as a coronary artery, withatherosclerotic plaque is appropriately sedated and,placed upon a x-rayor fluoroscopic table. Various know steps could be used for locating thetip 56 of laser catheter 14 at the site for treatment of the patient'sartery and only a basic discussion of the procedure for locating the tip56 at the proper site will be presented herein. Initially, the guidewire 20 and a guide catheter of common design (not shown) would beinserted into the patient using an introducer sheath of common design(not shown). The guide catheter would extend to the mouth of the artery,whereas the guide wire would be manipulated to anchor its end justbeyond the partial obstruction caused by the plaque.

The treatment catheter 10 (refer to FIG. 1), which includes both thelaser or inner catheter 14 and the insertion or flush catheter 12 wouldthen be slid along the guide wire 20. The catheters 12 and 14 would movealong the guide catheter (not shown) in known fashion until the balloon16 of the insertion catheter 12 is outside of the guide catheter anduntil the window portion 56W is adjacent the portion of the artery forwhich treatment is intended. The laser catheter 14 would be rotateduntil its window portion 56W (refer to FIG. 2) faces the side of theartery wall which is to be treated. The balloon 16 is then inflatedusing the balloon syringe 22 such that the part of the artery downstream(it would correspond to the rightward direction in FIG. 1) of balloon 16is blocked from receiving further blood. The balloon 16 may be used toblock the blood flow for up to about 60 seconds. The blockage wouldnormally not need to be maintained for 60 seconds, but the surgeon wouldbe using his professional judgement as to how long the blockage might betolerated for a particular individual. At any rate, the blockage wouldbe less than two minutes at a time and is significantly less than theblockage times required for the double balloon prior art techniquedescribed in the background portion of this application. If advisable,the balloon could be deflated and re-inflated to provide repeatedtreatments without maintaining the blockage for longer than about 60seconds each time.

After the balloon 16 has been inflated and now considering FIGS. 1 and2, a saline or other flushing solution is supplied to connector 26. Thesaline travels along the flushing solution channel 48 and exits from 48E(see especially FIG. 2) so as to clean out blood in the portion of theartery just downstream from the balloon 16. After this space has beenflushed with saline, the saline flush is halted and the laser 40 isactivated to generate the laser energy 18 (FIG. 1) in the form of themicrobeams 64 (FIG. 2). The laser 40 would preferably be a pulsed laserpulsed at one to five times a second, such as a 355 nanometer tripledYAG or a flash lamp excited dye laser at 504 nanometers. However, acontinuous wave argon laser or other type of laser might be used. Atreatment agent will be supplied to the cells which are porated by thelaser microbeams. A treatment agent as used herein is a cell treatmentagent, meaning that it has medicinal effect (might include killing thecell if that was medically helpful) or harmful effect (if desirable fortesting purposes) or remedial effect when placed within a cell afterpassing through porations caused by the laser microbeams. The treatmentagent, such as genetic material or a drug is supplied to the connector30 (FIG. 1) and passes through the treatment agent channel 53 andthrough the space between optical fiber 38 and connecting tube 54 intothe hole or bore 56B for passage as a high pressure stream out of thewindow portion 56W. Referring now to FIG. 5, two cells 68 are shownhaving porations 68P therein as caused by the laser microbeams 64 havingedges 64A shown in FIG. 5, the microbeams having passed out of holes56H. Also passing out of the holes 56H is a high pressure solutioncontaining the treatment agent 70 disposed therein and some of thetreatment agent 70A has entered into the cells 68 by way of theporations 68P. If the agent 70 includes genetic materials, the solutionmay be culture material such as DMEM (Dulbecco's Modified Eagle'sMedium) or normal saline. Quite importantly, the treatment agent 70passes out of the same holes as the beams 64 such that the treatmentagent passes along the same paths as the various microbeams 64.Accordingly, the genetic material should enter through the porations 68Pin the cell walls of cells 68. In other words, the treatment agent isconcentrated precisely where it is most likely to be effective.Significantly, the beam width is smaller than the size (i.e., longestdimension) of the cell and would also preferably be smaller than thenormal dimension of the cell (i.e., the dimension of the cell extendingperpendicular to the direction of the beam).

It should be appreciated that some of the microbeams 64 may hit thenucleus of a cell and kill the cell. Others of the microbeams may hitthe edge of a cell without providing a useful poration. However, using alarge number of the microbeams should allow for treatment, on a cellularlevel, of a sufficient number of cells that benefits will be obtained.

As an alternative to the simultaneous spraying of treatment agent 70 outof the holes 56H while the beams 64 are passing out of the holes, onemight porate the cells 68 by application of the beams 64 and,immediately after turning off the beams 64, spray the treatment agent 70out of the holes 56H. Since the porations 68P will close relativelyquickly, the treatment agent should be sprayed immediately after turnoff of the beams.

If one is simply treating a single part of the artery wall, one mightline up the window portion 56W to face the proper direction by use of amarker (not shown) on part of the laser catheter 14. For example, a holeor pattern (not shown) might be placed on the side of steel tip 56opposite to the window portion 56W. The surgeon would then observe themarker by use of the x-ray table and rotate the laser catheter 14 untilthe marker was opposite to the part of the artery wall which was to betreated. If desired, two markers of different configuration might beused to provide more information to the surgeon and to help better lineup the window portion 56W such that it faces the part of the artery wallwhich is to be treated. Instead of placing the markers on the steel tip56, the markers might alternately be placed upon the tapered portion 58(which is made of plastic) and/or the outer surface of the lasercatheter 14. If desired, one may treat the artery wall in a completecircumference. One may apply laser energy and treatment agent (eithersimultaneously or treatment agent immediately after laser as discussedabove) with the laser catheter 14 disposed in one angular position. Thelaser would then be turned off, the laser catheter 14 would be rotatedto a different angular position and the laser and treatment agentapplication would be repeated. The laser would be turned off and thelaser catheter 14 would be rotated to another angular position fortreatment. This process of rotation treatment followed by furtherrotation and treatment may be performed around the completecircumference of a portion of the artery. Additionally, or alternately,one may move the laser catheter 14 along the guide wire 20 to adifferent place within the artery before applying further treatment. Inother words, if the blockage or other problem in the artery extendssignificantly in a lengthwise direction, treatments may be applied atdifferent places along the length of the blockage.

Advantageously, a vacuum may be applied to tube 28 to remove geneticmaterial or drug remaining free after laser operation and beforedeflating balloon 16. This reduces the amount of material going downstream.

After application of the laser beams and treatment agent has beencompleted, the balloon 16 would be deflated so as to reopen the artery.The treatment catheter 10 composed of laser catheter 14 and insertioncatheter 12, is then removed from the patient. The guide catheter andguide wire would be removed from the patient and normal post-operativeprocedures would be followed such as checking the patient to insure thatno artery walls were punctured.

Having shown how the present invention may be used to very efficientlyprovide treatment at a cellular level by injecting cell treatment agents70 directly into cells 68 as illustrated in FIG. 5, some specificexamples of such treatments using laser energy will now be presented.Generally, any genetic code material, such as DNA plasmids, and any drugapplicable for cellular treatment could be used.

EXAMPLE 1

A patient has a buildup of plaque on the walls of a coronary artery. Thetreatment catheter 10 would be inserted into the patient under theprocedure explained above and the microbeams 64 are used to poratesmooth muscle cells of the artery walls. The treatment agent would beplasmids of DNA which encode antisense gene. The gene may be under thecontrol of mouse metallothionein promotor. A virus carrier would be usedin known fashion to allow the desired genetic material to enter thenucleus and/or cytoplasm (most frequently the cytoplasm) of the cellsinto which the treatment agent is inserted. The antisense genetic codewill fool the smooth muscle cells to inhibit growth patterns which causethe blockage of arteries.

EXAMPLE 2

A patient has a buildup of plaque on the coronary arteries. Thetreatment catheter 10 would be inserted into the patient under theprocedure described above and the microbeams 64 are used to porate theendothelial cells of the artery walls. The treatment agent would beplasmids of DNA which encode a tissue plasminogen activator gene underthe control of the mouse promotor as with example 1 and having a viruscarrier. The human plasminogen activator would cause the production ofenzymes which reduce formation of clots.

EXAMPLE 3

A patient has a malignant tumor in the colon. The patient would besedated and a colonoscopy would be performed. Instead of using an x-rayor fluoroscopic table to locate the position of the treatment deviceplaced within the patient, the treatment device (not shown) may includean optical fiber to allow the surgeon to see within the colon. The probeor medical device inserted into the patient would include a lasercatheter similar to catheter 14 of FIG. 2. Upon the window portion(similar to 56W of FIG. 2) being lined up to face the tumor, laserenergy is applied to provide microbeams which porate the cells of thetumor and a cancer agent, such as 5-fluorouracil or donarubicin, isinjected out the same plurality of holes used for generating themicrobeams. Saline or other solution may be used to carry the canceragent drug to the cells. The cancer agent would enter into numerous ofthe cancer cells and kill them. Advantageously, the poration of thecells by the laser beam improves the efficiency of application of thecancer agent to the cells. A smaller portion of the cancer agent harmsadjacent healthy cells than would be the case if one simply applied thecancer agent against cells which had not been porated. Some of thecancer cells may be killed simply by application of the laser beam, butthe inclusion of the cancer agent helps to kill a greater portion of thecancer cells than would otherwise be the case. Additionally, if onesimply relied upon the laser to kill cancer cells, one might have to usea higher laser power which in turn might damage healthy cells behind oradjacent to the cancer cells.

EXAMPLE 4

The patient would be the same as in example 2 and the same procedurewould be followed except that the treatment agent is the drug heparincarried by saline or other solution. The drug prevents formation ofclots.

EXAMPLE 5

An animal may be used to test various anti-plaque techniques by usingthe present invention to induce plaque in walls of arteries. Thetreatment catheter 10 is used to porate endothelial cells on arterywalls of the animal (patient) for introducing a plasmid of DNA whichencodes for human growth hormone gene under the control of a mousemetallothionein promoter. A viral carrier would be used in known fashionto allow the desired genetic material to enter the nucleus and/orcytoplasm of the cells into which the treatment agent is inserted. Theexpression of growth hormone in transfected cells will then result inthe expression of various cellular proteins causing cell growth whichcould be responsible in part for the development of plaque in thearterial wall. This information could then be used to develop eitherdrugs or other methods to inhibit gene expression in order to block thisgrowth.

Turning now to FIG. 6, an alternate embodiment constructed in somewhatsimilar fashion to the embodiment of FIG. 1 will be discussed. Thecomponents of the FIG. 6 embodiment have numbers in the 100 series withthe same last two digits as the corresponding component, if any, of theFIG. 1 embodiment. Thus, the components 110 through 144 are identical inconstruction and operation as the corresponding components in FIG. 1except as discussed hereafter.

As shown, a reservoir 172 of treatment agent with DNA plasmid isconnected to pump 174. The pump 174 pumps the agent into entry tube 132by way of tube 176. This operation as described so far is identical tothe catheter 10 of FIG. 1 where the reservoir and pump were simply notshown. What is different about the catheter system 110 is the use of anelectrical heating coil 178 wrapped around the tube 176 in order to heatthe agent prior to its insertion in the patient. By heating the agentsufficiently that the agent adjacent the patient's cells will be from 42degrees to 45 degrees, the transfection process is speeded up and willoccur faster than would otherwise be the case. Indeed, the laser 140 andlaser catheter 114 could be left out of the apparatus 110 and the heatfrom electrical heater 178 could be used without laser energy in orderto expedite the transfection process. In that case, a catheter 114 couldbe constructed like catheter 14 of FIG. 2 except that there would be nooptical fiber such as 38 of FIG. 2. Also, such an arrangement could useone of more large exit Holes instead of the array of 50 to 100 microndiameter holes 56H of FIG. 4.

Although the heater 178 is shown as a coil heater, other electrical ornon-electric heating techniques could be used to heat the agent eitherprior to its insertion in the catheter 110 or while it is travelingtherein.

FIG. 7 shows an embodiment using application of laser energy to heat thetip of catheter 210 and thereby indirectly heat the cells. Thecomponents of the FIG. 7 embodiment have numbers in the 200 series withthe same last two digits as the corresponding component, if any, of theFIG. 1 embodiment. Insertion catheter 212, laser catheter 214, opticalfiber 238, flushing solution channel 248, flushing solution exit 248E,and treatment agent channel 253 function as the corresponding componentsin the previous embodiments except as discussed below. (The unseen partsof catheter 210 would be connected to a laser and a source of treatmentagent with DNA plasmid as with previous embodiments.)

The FIG. 7 embodiment is different from the others in that it has anopaque metal cap 280 with neck portion 280N crimped or otherwise fixedto catheter 214. The metal may be similar to metal parts 50 in FIG. 4and 70 in FIG. 5 of the present inventor's U.S. Pat. No. 5,041,109,issued Aug. 20, 1991, and hereby incorporated by reference in itsentirety. Cap 280 has holes 280H for treatment agent to exit from,whereas optic fiber 238 terminates in a spherical lens 238S whichdistributes laser energy, not shown, over a wide area of cap 280. Arms238A may be circumferentially arranged to support fiber 238. The laserenergy heats the cap 280, which in turn would heat the patient's cells(not shown) and make them more receptive to transfection from thetreatment agent.

Although not shown in the FIG. 7 embodiment, one or more occluderballoons such as 16 of FIG. 1 could be used in the FIG. 7 embodiment andthe embodiments hereafter discussed. For ease of illustration, theinsertion catheter, such as 212 of FIG. 7, and flushing solution channelwill not be shown or discussed for the embodiments discussed below, eventhough those embodiments would have such features.

Catheter 310 of FIG. 8 has numbers in the 300 series with the same lasttwo digits as the corresponding component, if any, of the earlierembodiments. This embodiment has a cap 380 with hole 380H for dispersingtreatment agent from channel 353. A thermocouple or other temperaturesensing device 381 has wires 381W (only one shown for ease ofillustration) extending out of catheter 310. The cap 380 has at leastone heating resistor 382 embedded therein or otherwise in thermalcontact therewith. Wires 382W (only one shown for ease of illustration)carry electrical current from a power supply 382 external outside of thepatient to the heater 382. The power supply 382P is part of a feedbackcontrol closed loop which uses negative feedback by way of temperatureadjustor resistor 381R and comparator 381C. As well known such a controlloop could alternately use an amplifier, subtractor, or adder (insteadof comparator 381C) to generate a difference signal at the outputthereof to cause the heater 382 to heat more if the temperature is lowand to heat less if the temperature is too high. Since thermocouple orother sensor 381 is located near the hole 380, it will very accuratelytrack the temperature of treatment agent fluid passing out the hole380H. Preferably, the sensor 381 is within at least 5 inches of hole380H, and more preferably and advantageously within 3 inches thereof.Most preferably, it is at the edge of hole 380H or within 1 inch of theedge of the hole.

The feedback circuit may be configured differently from FIG. 8 as manytypes of thermostatic or other feedback techniques are known forstabilizing a sensed condition such as temperature. It should beemphasized that the feedback control to maintain the fluid temperatureshown for FIG. 8 would preferably be used for the various otherembodiments discussed above and below. Such feedback control loops andcomponents are not shown for the other embodiments for ease ofillustration. However, all embodiments may use a temperature dependentsensor on the catheter as discussed to control the output of the heatingdevice or devices so as to maintain the temperature at a desired valueor within a desired range, which is preferably 42 to 45 degreescentigrade.

The heater 382 may be of the CAL-ROD type or any other device generatingheat from electricity. The cap 380 may be metal like cap 280 or could beother material suitable to disperse heat to patient's cells adjacent tothe window 380H. As with the other embodiments not based on directapplication of laser energy to the patient's cells, the heat could beused to heat the patient's cells to from 42 to 45 degrees centigrade andenhance their receptivity to transfection.

Catheter 410 of FIG. 9 has numbers in the 400 series with the same lasttwo digits as the corresponding component, if any, of the earlierembodiments. Inner catheter 414 has holes 480H to allow exit oftreatment agent from treatment agent-channel 353. Radio frequency (RF)waves are emitted from antenna 484 or other known RF source. Wires 484Ware connected to supply power as appropriate to antenna 484, which isembedded in part 414E. Part 414E may simply be a generally cylindricalrounded tip of the material making up the side of catheter 414 (andsides of the various other catheter embodiments), which material may bePTFE (such as Teflon) or other materials commonly used for catheters.The RF energy is used in this embodiment to heat the patient's cells,preferably to 42 to 45 degrees centigrade, in order to make them morereceptive to transfection.

The FIG. 10 embodiment of catheter 510 uses microwave energy from source586 outside the patient to heat cells for improving receptivity totransfection. Microwave source 586 supplies microwave energy to innercatheter 514 by way of waveguide 586G for exiting at tip 586T. Themicrowave energy is applied to patient's cells (not shown) adjacent hole580H from which treatment agent from channel 553 may exit.

FIG. 11 shows catheter 610 with inner catheter 614 having an infraredsource 688 connected by wires 688W to a power source (not shown) outsidethe patient. Infrared energy from source 688 heats treatment fluidand/or the patient's cells (not shown) as treatment agent from channel653 exits hole 680H.

The catheter 710 of FIG. 12 has an inner catheter 714 with a treatmentagent channel 753 supplying treatment agent out hole 780H. In thisembodiment a chemical compartment behind (right in FIG. 12) wall 790 isdivided into top and bottom parts 790T and 790B by separation wall 790S.Parts 790T and 790B hold different chemicals which react to release heatwhen they mix upon control line 790C being used to slide separation wall790S leftwardly in FIG. 12. The heat heats patient cells to improvereceptivity to transfection.

FIG. 13 shows a catheter 810 with inner catheter 814 having hole 880Hfor releasing treatment agent to a site 792S within a patient's artery(or other body passage) 792A. In this arrangement the cells at site 792Sare heated by application of ultrasound energy from transducers 794 onthe skin 792K of the patient.

Turning now to FIGS. 14-17, a further embodiment catheter 910 has a tip956 with tapered portion 958. A guide wire 920 is used and a glass hood960 is used. The components of catheter 910 and other unshown componentsmay be constructed and may operate as the corresponding component (samelast two digits) from FIGS. 1-6. This discussion will proceed withdifferences between catheter 910 and catheter 10.

Catheter 910 has a window 956W which is simply one large opening.Instead of relying on holes to produce microbeams of laser energy forcell poration as done in the embodiment of FIGS. 1-6, catheter 910 usesa bundle 995 having a sloped tip 995S and a plurality of optical fibers938 (only some numbered for use of illustration) with filler material995C. At least the portions adjacent to slope 995S of the fillermaterial should be clear such that beams 964 may emerge from the tips938T (shown schematically in FIG. 14 only). Each tip 938T may provide asingle corresponding beam 964, which beam may have dimensions andoperate as discussed for each of the beams 64 of FIG. 2. As illustratedschematically in the front view of FIG. 14, the tips 938T and fibers 938are staggered such that light from one tip 938T will not hit another ofthe tips 938T or fibers 938 on its way to the single opening window 956Wand the patient tissue outside it.

The laser energy at the tips 938T is reflected out the window 956W byeither of two arrangements. A first way is to simply have the opticalfiber tips 938T cut at the same angle (preferably between 35 and 55degrees, most preferably an angle from 42 to 48 degrees, with 45 degreesbeing the most preferred value) as the slope 995S. (The angle beingmeasured relative to a horizontal line, not shown, perpendicular to theaxes of the parallel optical fibers 938.) This first way has tips whichreflect light such that it goes in the direction from a low end of thesloped face towards the high end of the sloped fiber tip. A second wayis to have a prism such as shown and explained for the tip of FIG. 7 ofthe incorporated by reference U.S. Pat. No. 5,061,265.

FIG. 17 shows the external end 995E of bundle 995 receiving a laser beam940B from a laser 940. Since the optical fibers are relatively small,they will produce a plurality of beams having the width ranges asdiscussed with respect to the embodiment of FIGS. 1-6. The opticalfibers themselves may have diameters equal to the various rangesdiscussed with respect to the beam widths given above dimensions withrespect to the embodiment of FIGS. 1-6. The optical fibers may be 80microns in diameter. As in the other drawings, only a relatively smallnumber of fibers 938 are shown, but it will be understood that thenumber of fibers could be just a few or a relatively large number.

A further alternate to the arrangement of FIGS. 14-17 may be discussedwith reference to FIG. 16 since such a further embodiment might beconstructed as with FIGS. 14-17 and have essentially the same top view.The beam pattern of FIG. 16 might be realized by plurality of fiberswith prisms at their tips and with all the tips ending in the same planeperpendicular to the axes of the fibers and the catheter. Further, suchprisms could alternately direct the beams radially outward from acentral axis of a bundle such as 995S.

Turning now to FIGS. 18 and 19, catheter 1010 has a tip 1056 and bundle1095 operating and constructed as with the FIG. 15 embodiment except asfollows. The bundle 1095 has a flat tip end 1095T from which beams 1038Bpass from a plurality of fibers 1038 to a reflector 1096, which may beeither a piece of silicon glass or a mirrored reflector. The reflector1096 may be made and operate in the fashion explained in Saadatmanesh etal. U.S. Pat. No. 5,242,438, issued Sep. 7, 1993 and hereby incorporatedby reference except that the present reflector is shaped differently asdiscussed hereafter.

The reflector 1096 has a series of parallel ridges 1096R and valleys1096V extending across reflector 1096 perpendicular to the plane of viewof FIGS. 18 and 19. The ridges and valleys are stepped in that thoseclosest the one-hole window 1056W are further from tip end 1095T thanthe ridges and valleys which are further from the window 1056W. Sincethe slopes between ridges 1096R and valleys 1096V which face the window1056W have a 45 degree slope relative to beams 1038 (corresponding tothe axes of fibers 1038), beams 1038 are reflected at right angles (90degrees) becoming beams 1064. The beam widths for beams 1064 would be asdiscussed with respect to FIGS. 1-6.

FIG. 20 shows a bottom view (i.e., side receiving laser energy) of analternate reflector 1196 similar in operation and construction asreflector 1096 except as follows. Reflector 1196 has a plurality of rows1197 of width W, each row having stepped ridges 1196R and valleys 1196Vsuch that a cross section along or parallel to any of lines 1197R (whichseparate adjacent rows) reveals a profile like that presented byreflector 1096 in FIG. 19. By using a plurality of the rows 1197, asingle optical fiber like fiber 38 of FIG. 2 (but without a prism at itstip) could apply a relatively wide beam to reflector 1196. The reflector1196 would receive such a single beam (not shown) exiting in line withthe central axis of the single optical fiber and create multiple beamsby reflection. The reflection would be accomplished as discussed forFIG. 19 except that the reflection would actually be separating arelatively wide beam into a plurality of beams (not shown). The beamwidths would be as discussed with respect to FIGS. 1-6.

It will be readily appreciated that the embodiments of FIGS. 7 to 20would be constructed in the same fashion as the FIGS. 1 to 6 embodimentsexcept for the illustrated differences. All of the embodiments whichheat the patient's cells in vivo without direct application of laserenergy to the cells would heat the cells to 42 to 45 degrees centigradefor improving transfection receptivity. All of the embodiments may useone or two occluder balloons as discussed in connection with the FIG. 1embodiment and may use feedback control as discussed with respect toFIG. 8. Although the discussion of FIGS. 6 to 20 has concentrated on DNAplasmids for treatment, drugs or other materials could be used. Thetreatment agent may have plasmids coprecipitated with CaPO₄ or someother catalyst as that may be at least advisable to improvetransfection.

Tests by the present inventor of in vitro transfection into bovine aortasmooth muscle cells (SMC) have shown that heat can increase theefficiency of transfection. The plasmid pXGH5, which encodes a humangrowth hormone (hGH) reporter, was used to determine efficiency oftransfection. SMC transfection was performed using DNA coprecipitatedwith CaPO₄. SMCs were immediately heated for up to 1 hour at 42 to 45degrees centigrade. Results were compared to paired control experimentsconducted on the same day in the absence of heating. Transient geneexpression was determined by radioimmunoassay of spent medium for hGH 48hours after transfection. SMCs heated and exposed to DNA without CaPO₄showed no gene expression. In the 60 paired experiments, SMCstransfected by CaPO₄ coprecipitation of DNA and heated demonstrated anincreased hGH production compared to unheated controls, 20.3 plus orminus 3.7 versus 16.8 plus or minus 3.4 (mean plus or minus SE)respectively, p<0.002.

With respect to the various embodiments applying laser energy to thetissues of a patient, one may use an external chromophore which willabsorb wavelengths selectively to raise the temperature of the desiredtissue. This would selectively heat the target tissue and reduce heatapplied to other tissues. Also, it should smooth out the process ofheating (i.e., avoid too fast temperature changes) and provide a moreeven temperature distribution over the desired tissues. Such achromophore may involve oral or intravenous injection of materials whichwill concentrate in the desired tissues and make them more receptive tolaser heating. Also, such materials to increase receptivity to laserheating may be inserted in the channel or channels of the catheter to bedirectly applied adjacent the heating.

Although specific constructions and examples have been presented herein,it is to be understood that these are for illustrative purposes only.Various modifications and adaptations will be apparent to those of skillin the art. For example, one might use a double balloon catheter withthe laser beams and treatment agents being applied from a window portionin between two balloons. Although the present invention has the highlyadvantageous feature of injecting the treatment agents out the sameholes as the laser beams, the present invention, in its broadestaspects, might include a double balloon arrangement wherein thetreatment agent comes out holes separate from the laser beams and fillsthe chamber established between the two balloons blocking part of anartery. Since the laser beams would be porating the cells within arterywalls between two such balloons, the treatment agent, such as geneticmaterial, could transfect the cells more quickly than in the prior artdouble balloon technique discussed in the background portion of thisapplication since that prior art technique did not provide for cellporations. Such a double balloon technique may also use known dyematerials inserted to enhance absorption of laser energy. Such materialsare disclosed in the present inventors' prior U.S. Pat. Nos. 4,860,743and 5,041,109 issued respectively on Aug. 29, 1989 and Aug. 20, 1991 andhereby incorporated by reference. Although the laser 40 would preferablybe a pulsed type laser which could use feedback control as discussed,one would especially want to use a thermocouple (for feedback control asdiscussed) to guard against overheating if the laser 40 was a continuouswave laser. In view of these and other possible modifications, it willbe appreciated that the scope of the present invention should bedetermined by reference to the claims appended hereto.

What is claimed is:
 1. A system for applying treatment fluid within apatient at a desired temperature comprising: a catheter having a fluidchannel and fluid port therein; a fluid source operably connected to thefluid channel; a heater for fluid passing within the fluid channel, theheater being operably connected to the fluid channel to control the heatof fluid in at least part of the fluid channel; a temperature sensor onsaid catheter positioned to sense fluid temperature adjacent said fluidport; and a closed feedback control loop connecting said temperaturesensor to said heater and operable to automatically stabilize thetemperature of fluid passing out said fluid port, the closed feedbackcontrol loop including a means to compare a sensed temperature with adesired temperature and generate a signal controlling power output fromthe heater; and wherein said temperature sensor is on a portion of saidcatheter, which catheter portion is inserted in a patient during use;and further comprising: a source of treatment agent selected from thegroup consisting of genetic material and drugs, operatively connected tothe fluid channel so as to apply treatment agent to heated cells by wayof the fluid channel; and wherein the treatment agent is operable totreat the heated cells, the heater being operable to heat the treatmentagent such that treatment agent effects on the heated cells areenhanced.
 2. The system of claim 1 wherein said heater generates itsheat at said catheter portion.
 3. The system of claim 1 wherein saidheater is in said catheter portion.
 4. The system of claim 1 whereinsaid heater is an electrical resistance heater.
 5. The system of claim 1wherein said heater is an RF energy heater.
 6. The system of claim 1wherein said heater is a microwave heater.
 7. The system of claim 1wherein said heater is an infrared heater.
 8. The system of claim 1wherein said heater is a chemical reaction heater generating heat by achemical reaction in said catheter portion.
 9. The system of claim 1wherein said heater includes a laser.
 10. The system of claim 9 whereinsaid heater includes an optical fiber conveying laser energy from saidlaser to said catheter portion.
 11. The system of claim 1 wherein saidheater is one or more ultrasound transducers operable to apply heat toskin of the patient such that the heat may pass to the catheter portion.12. The system of claim 1 wherein the heater is operable to heat cellsof the patient in vivo and further comprising: a source of treatmentagent including a DNA plasmid operatively connected to the fluid channelso as to apply treatment agent to the heated cells by way of the fluidchannel; and wherein the treatment agent is operable to transfect theheated cells.
 13. A system for applying fluid within a patient at adesired temperature comprising: a catheter having a fluid channel andfluid port therein, the catheter having a catheter portion, whichcatheter portion is inserted in a patient during use; a fluid sourceoperably connected to the fluid channel; a heater for fluid passingwithin the fluid channel, the heater having at least a portion mountedto the catheter, the heater being operable to control the heat of fluidin at least part of the fluid channel; a temperature sensor on saidcatheter positioned to sense fluid temperature adjacent said fluid port;and a closed feedback control loop connecting said temperature sensor tosaid heater and operable to automatically stabilize the temperature offluid passing out said fluid port, the closed feedback control loopincluding a means to compare a sensed temperature with a desiredtemperature and generate a signal controlling power output from theheater; and wherein said temperature sensor is on the catheter portion;and further comprising: a source of treatment agent selected from thegroup consisting of genetic material and drugs, operatively connected tothe fluid channel so as to apply treatment agent to heated cells by wayof the fluid channel; and wherein the treatment agent is operable totreat the heated cells, the heater being operable to heat the treatmentagent such that treatment agent effects on the heated cells areenhanced.
 14. The system of claim 13 wherein said heater generates itsheat at said catheter portion.
 15. The system of claim 13 wherein saidheater is in said catheter portion.
 16. The system of claim 13 whereinsaid heater is an electrical resistance heater.
 17. The system of claim13 wherein said heater is an RF energy heater.
 18. The system of claim13 wherein said heater is a microwave heater.
 19. The system of claim 13wherein said heater is an infrared heater.
 20. The system of claim 13wherein said heater is a chemical reaction heater generating heat by achemical reaction in said catheter portion.
 21. The system of claim 13wherein said heater includes a laser.
 22. The system of claim 21 whereinsaid heater includes an optical fiber conveying laser energy from saidlaser to said catheter portion.
 23. The system of claim 22 wherein theheater is operable to heat cells of the patient in vivo and furthercomprising: a source of treatment agent including a DNA plasmidoperatively connected to the fluid channel so as to apply treatmentagent to the heated cells by way of the fluid channel; and wherein thetreatment agent is operable to transfect the heated cells.